Adaptive thermogenesis is the cellular process to generate heat in response to environmental stimuli such as low temperature, which requires the conversion of white adipocytes into UCP1-expressing beige adipocytes (WAT browning). The increase of adaptive thermogenesis is an attractive target to counteract obesity and metabolic diseases. However, contribution of dietary components to adaptive thermogenesis is largely unknown. The long-term goal of Dr. Soonkyu Chung?s laboratory is to develop efficacious and safe dietary strategies that can help control human obesity by increasing adaptive thermogenesis. The laboratory?s recent preliminary data suggest that palmitate reduce UCP1 expression, which was normalized by DHA treatment in human beige adipocytes. The aim of COBRE Research Project 5 is to identify the underlying mechanisms by which n-3 PUFA enhance adaptive thermogenesis by promoting beige conversion. Dr. Chung hypothesizes that inhibition of TLR4 signaling by n-3 PUFA will antagonize innate immune responses by SFA and subsequent autophagic mitochondria degradation, leading to enhanced WAT browning and thermogenesis. To test this hypothesis she proposes two specific aims. In Specific Aim 1, she proposes to identify the early signaling crosstalk by which SFA versus n-3 PUFA differently modulate white-to-beige adipocyte transition in human beige adipocytes. She will determine different dietary fatty acids (FA) regulate TLR4 activation, autophagic degrading, and mitochondria dynamics using the primary human beige adipocytes that she has recently established from multi-potent human adipose-derived stem cells. In Specific Aim 2, Dr. Chung will determine the impact of TLR4 signaling deficiency on the thermogenic capacity and metabolic consequences in vivo. Inhibition of TLR4 signaling will be achieved by dietary alleviation with n-3 PUFA and by genetic deletion of TLR4/MyD88 signaling in obesity prone C57BL/6 mice. The proposed study fits well with the Nebraska Center for Prevention of Obesity Diseases through Dietary Molecules (NPOD)?s thematic focus because it strives to identify the novel signaling pathways by which dietary FA regulate energy metabolism and innate immunity. Successful completion of the proposed study will provide the first evidence that dietary FA composition (degree of desaturation and n-6/n-3 ratio) differentially modulates adaptive thermogenesis. This information will be significant for devising successful dietary strategies to increase the adaptive thermogenesis.